Isobenzofuran analogs of sclerophytin A

ABSTRACT

Isobenzofuran analogs of sclerophytin A are prepared in a highly concise fashion via an aldol-cycloaldol sequence. The analogs exhibit IC 50 &#39;s as low as 1 μM in growth inhibitory studies against KB3 cells using an MTT assay. Preferred analogs have one of the following structural formulas, where R is hydrogen or a substituted or unsubstituted lower alkyl group and Ar is a substituted or unsubstituted aryl group.

REFERENCE TO RELATED APPLICATION

The present application claims the benefit of U.S. ProvisionalApplication No. 61/269,875, filed Jun. 30, 2009, the disclosure of whichis incorporated by reference herein.

STATEMENT OF GOVERNMENT SUPPORT

The present invention has been supported in part by NIH Grants CA75577,RR15569, CA125602 and CA109821. The Government has certain rights in theinvention.

FIELD OF THE INVENTION

The present invention relates to isobenzofurans, particularly compoundsstructurally related to sclerophytin A, having anti-cancer and/orantibiotic activities.

BACKGROUND OF THE INVENTION

The 2,11-cyclized cembranoids are a class of diterpenoids isolated froma variety of marine sources that display a range of biologicalactivities.(1) Some 2,11-cyclized cembranoids reported to possesscytotoxic activity are shown below:

Sclerophytin A has been reported to exhibit growth inhibitory activityagainst the L1210 cell line with an IC₅₀ of 1.0 ng/mL (3 nM).(2-4)Eleutherobin(5,6) and sarcodictyins(7,8) A and B are reported to exhibittaxol-like anti-mitotic activity.(9,10) There have been several reportsof structure/activity relationship studies of sarcodictyin (11,12) oreleutherobin(13-16) analogs. However, there are fewer biological studiesof any analogs of the more common isobenzofuran-containing 2,11-cyclizedcembranoids.(17,18)

Recently, synthetic approaches have been developed for making selectedcyclized cembranoids in which the isobenzofuran bicycle is assembled inas few as three steps from commercially available (S)-(+)-carvone usinga highly stereoselective aldol-cycloaldol sequence.(19) This approach isused herein to prepare structurally related isobenzofurans that arenominal analogs of sclerophytin A.

SUMMARY OF THE INVENTION

The present invention is directed to novel isobenzofuran analogs ofsclerophytin A, which have anti-cancer and/or antibiotic properties.Some preferred compounds of the present invention have one of thefollowing basic structural formulas:

In the formulas, R represents H, or a lower alkyl group, which issubstituted or unsubstituted, and Ar represents a substituted orunsubstituted aryl group. A particularly preferred compound has formula10, wherein Ar is 2-fluorophenyl (herein referred to as compound 10h). Acompound of the present invention can have a structure indicated above,or can be provided as a pharmaceutically acceptable ester, salt, orprodrug that releases a compound of the invention when metabolized.

Also contemplated is a method of treating a patient suffering fromcancer or a microbial infection comprising administering to the patienta therapeutically effective amount, i.e., one that inhibitsproliferation of the cancer or microbial cells, of a compound of thepresent invention.

BRIEF DESCRIPTION OF THE DRAWING

FIGS. 1A-D show the results of inhibition studies of KB-3 cell viabilityfor some selected isobenzofuran compounds (Panel 1A: 5a; Panel 1B: 5c;Panel 1C: 6a; Panel 1D: 8). Cells were treated with increasingconcentration of compounds and MTT viability assays were performed after96 hours. Values are the means of triplicate assays and are expressed asmean±standard deviation relative to control (untreated cells). Note—theX-axis has a non-linear scale.

DETAILED DESCRIPTION OF THE INVENTION

A compound of the present invention has a chemical structure selectedfrom the following formulas:

In the formulas, R represents H, a substituted lower alkyl group, or anunsubstituted lower alkyl group. Ar represents a substituted orunsubstituted aryl group. Pharmaceutically acceptable esters, salts, andprodrugs of such compounds are also contemplated within the scope of theinvention.

Preferably, in the formulas R=methyl, ethyl, cyclopropylmethyl orcyclopentylmethyl. It is also preferred that Ar=2-Br-phenyl,3-Br-phenyl, 2,3-di-Cl-phenyl, 2,4-di-Cl-phenyl, 1-naphthyl, 2-pyridyl,2-furyl, or 2-fluoropheynyl. A particularly preferred compound hasstructure 10, wherein Ar=2-fluorophenyl.

A method of synthesizing a compound of the present invention comprisesconverting (S)-(+)-carvone to an aryl glycolate derivative thereof. Theconversion comprises reacting (S)-(+)-carvone with an arylaldehyde in analdol condensation reaction to afford an aryl anti-alcohol, andetherifying the aryl anti-alcohol to afford the aryl glycolatederivative. The aryl glycolate derivative can then be cyclized to affordan isobenzofuran having structural formula 5. The isobenzofuran can thenbe converted by oxidative rearrangement to an enone having structuralformula 6. Alternatively, the aryl glycolate derivative can be convertedby β-lactonization-decarboxylation to a diene having structural formula10.

Synthesis of isobenzofurans of the present invention bycycloaldolization is illustrated in Scheme 1. Intermolecular aldolreaction of (S)-(+)-carvone (1) and aryl aldehydes 2a-f gives anti-aldoladducts 3a-f in moderate to excellent yields and with diastereomerratios from 1-20:1. Etherification of alcohols 3a-f proceeds in moderateto high yields to give glycolate esters 4a-f. Cycloaldolization underthe influence of KHMDS afforded isobenzofurans 5a-f as singlediastereomers based on ¹H NMR analysis.

Sclerophytin A and all other 2,11-cyclized cembranoids possess a C10stereocenter in the alkane, rather than alcohol, oxidation state. Thereduction of alcohols like 5a-d in a 3-step process via thecorresponding enone have been previously reported.(19) Hence, enones6a-d were prepared by oxidative rearrangement of the corresponding 3°alcohols.(20)

Reduction of the C10 stereocenter is illustrated by the conversion ofenone 6a to tosyl hydrazone 7, which is followed by reductivetransposition to afford cis-fused isobenzofuran 8 (Scheme 2).

A series of acyl derivatives (shown below) of ester 5a were prepared toevaluate the effect of the carboxyl substituent on activity. Esters 5awere prepared by transesterification in the presence of excess alcoholand Bu₃SnOAc catalyst, while the corresponding carboxylic acid 9 wasprepared by saponification of ester 5a.

In the course of optimizing the cycloaldol reaction of glycolate 4a toester 5a, it was found that diene product 10 was formed in significantamounts (ca. 15%) when the reaction mixture is maintained at −78° C. forlonger periods of time prior to quenching with HOAc (Scheme 3). Formethyl glycolates 4g and 4h, the diene product was the only cyclizedproduct isolated from the reaction mixture. The dienes are presumablyformed by lactonization of the aldol intermediates to form unstableβ-lactones 11, which upon loss of CO₂ afford dienes 10.(21)

Finally, 2-pyridyl analog 5f was methylated to give pyridinium iodide 51(Scheme 4). Pyridinium salt 5i could serve as a cationic isostere of5.a.i.

Biological Assays

The human KB-3 carcinoma cell line was used to perform the MTTcolorimetric assay of cell viability.(22-24) Cells were treated withincreasing concentrations of compounds to assess their growth inhibitoryproperties. IC₅₀ values (concentration of the drug required to reducecell viability by 50%) ranged from 1 to >100 μM (Table 1). Theconcentration curve for 5c, which had an IC₅₀ of 5 μM, is shown in FIG.1, Panel B. Compounds 5a (FIG. 1, Panel A) and 6a (FIG. 1, Panel C) hadhigher IC₅₀ in the range of 20 and 70 μM, respectively. Several othercompounds, e.g., 8 (FIG. 1, Panel D), showed no evident growthinhibitory activity at the highest concentration tested (100 μM).

TABLE 1 Inhibition of KB-3 cell survival assessed by MTT viabilityassay. Entry Compound IC₅₀ (μM) 1  5a 20 2  5a.i 5 3  5a.ii 50 4 5a.iii >100 5  5b 3 6  5c 5 7  5d 3 8  5e 10 9  5f 100 10  5i >100 11 6a 70 12  6b 30 13  6c 5 14  6d 3 15  8 >100 16  9 >100 17 10a 4 1810e >100 19 10g 2 20 10h 1Discussion

Several structure-activity trends become apparent upon examination ofthe assay data. Firstly, for the 2-Br ester compounds 5a (entries 1-6),the smaller the alcohol moiety of the ester, the lower the IC₅₀, i.e.Me<Et<cyclopropylmethyl<cyclopentylmethyl˜neryl. Secondly, the ethylester alcohols 5a-d as a group were more active than the ethyl esterenones 6a-d as a group (entries 1, 5-7 vs. 11-14), with individualdifferences in the IC₅₀ values in the groups varying by factors of 1 to23.

The essentially equal potency of hydroxy esters 5b-d (entries 5-7) anddienes 10a, g, h (entries 17, 19, 20) is intriguing and suggests thatthe esters may undergo lactonization and decarboxylation to the dienesin vivo (cf. Scheme 3). This notion is supported by the substantialdifference in activity between β-OH ester 5a, which can undergolactonization, and the corresponding C10 reduction product 8, whichcannot (entries 1 and 15, respectively). On the other hand, the1-napthyl methyl ester 5e (entry 8) is at least 10-fold more active thanthe corresponding diene 10e (entry 18). Furthermore, enones 6a-d(entries 11-14) exhibit essentially equal potency to the β-OH methyl andethyl esters, and also cannot undergo β-lactone formation. Furtherstudies would be needed to determine whether β-hydroxy esters 5, enones6, and dienes 10 are inhibiting growth by the same or differentmechanisms of action.

The most active representatives of the compound classes (alcohol 5d,enone 6d and diene 10h) were submitted to NCI's DevelopmentalTherapeutics program for 60-cell line screening. Single dose assaysrevealed no significant activity for alcohol 5d. Enone 6d exhibitedsignificant differential activity against the RPMI-8226 leukemia and thePC-3 prostate cancer cell lines. Diene 10h was the most active compoundtested, possessing significant differential activity against the entireleukemia panel and the NCI-H522 non-small cell lung cancer cell line.Subsequent 5-dose testing of 10h revealed a GI₅₀=0.148 μM and LC₅₀=9.36μM for the RPMI-8226 leukemia cell line, and a GI₅₀=0.552 μM andLC₅₀=26.8 μM for the HOP-92 non-small cell lung cancer cell line.

CONCLUSION

A structurally novel set of analogs has been developed based on theisobenzofuran bicycle common to most of the 2,11-cyclized cembranoidsthat exhibit IC₅₀'s as low as 1 μM for growth inhibition against KB3cells. Analog 10h possesses sub-micromolar growth inhibitory activityagainst the RPMI-8226 leukemia and HOP-92 non-small cell lung cancercell lines.

The present invention is now described with reference to certainexamples, which explain but do not limit it.

EXAMPLES Example 1 Anti-alcohol 3a

To a solution of diisopropylamine (49.9 mmol, 1.5 eq) in dry THF (200mL) was added n-BuLi (49.9 mmol, 1.5 eq) dropwise at −78° C. under anitrogen atmosphere. After stirring for 15 min, (S)-(+)-carvone 1 (33.3mmol, 1 eq) was added drop-wise to the mixture at −78° C. After 30 min2-bromobenzaldehyde 2a (39.9 mmol, 1.2 eq) was added to the reactionmixture over about 45 min. The mixture was allowed to stir at −78° C.until no starting material was observed by TLC analysis (˜6 h). GlacialHOAc (49.9 mmol, 1.5 eq) was then added at −78° C. After stirring ˜5 minthe reaction was warmed to room temperature (rt) and diluted with 100 mLether, washed with water (150 mL) and extracted with ether (3×100 mL).The combined organic extracts were then washed with brine, dried overMgSO₄, and concentrated in vacuo. The crude product was purified viaflash chromatography eluting with 10:90 EtOAc/hexanes to deliver thedesired anti-alcohol 3a as a colorless oil (8.56 g, 77%). ¹H-NMR (270MHz, CDCl₃) δ 7.63-7.62 (d, 1H), 7.52-7.47 (q, 1H), 7.36-7.34 (m, 1H),7.14-7.11 (q, 1H), 6.70 (s, 1H), 5.22-5.18 (t, 1H), 4.81-4.73 (d, 2H),3.20-3.18 (d, 1H), 3.00-2.98 (t, 1H), 2.64-2.68 (m, 2H), 1.77 (s, 3H),1.67 (s, 3H). ¹³C-NMR (270 MHz, CDCl₃) δ 200.7, 145.1, 143.7, 141.3,135.3, 132.7, 129.3, 129.1, 127.6, 113.1, 72.5, 55.1, 44.3, 29.4, 20.9,15.8. IR (film) 3435, 2922, 2360, 1659 cm⁻¹.

Example 2

Anti-alcohol 3b. Following the protocol used for preparation of 3a thedesired alcohol 3b was obtained as a colorless oil in 75% (8.17 g, 25.13mmol) yield from 5g (33.22 mmol) (S)-(+)-carvone. ¹H-NMR (270 MHz,CDCl₃) δ 7.51 (s, 1H), 7.35-7.31 (m, 1H), 7.23-7.20 (t, 1H), 6.71-6.68(m, 1H), 4.91-4.89 (d, 2H), 4.81-4.76 (dd, 1H), 3.22 (d, 1H), 2.94-2.85(m, 2H), 2.40-2.43 (m, 2H), 1.74 (s, 3H), 1.53 (s, 3H). ¹³C-NMR (270MHz, 1DCl₃) δ 201.9, 144.8, 144.1, 142.6, 135.5, 134.5, 133.0, 127.6,127.1, 113.9, 70.4, 45.6, 31.3, 30.1, 20.6, 15.9

Example 3

Anti-alcohol 3c. Following the protocol used for preparation of 3a thedesired alcohol 3c was obtained as a colorless oil in 75% (8.17 g, 25.13mmol) yield from 5g (33.22 mmol) (S)-(+)-carvone. ¹H-NMR (270 MHz,CDCl₃) δ 7.66-7.63 (d, 1H), 7.38-7.36 (d, 1H), 7.28-7.25 (t, 1H),6.71-6.69 (m, 1H), 5.24-5.21 (dd, 1H), 4.65-4.60 (d, 2H), 3.16-3.10 (dd,1H), 3.07-2.95 (m, 2H), 2.46-2.40 (m, 2H), 1.84 (s, 3H), 1.39 (s, 3H).¹³C-NMR (270 MHz, CDCl₃) δ 201.9, 144.8, 144.1, 142.6, 135.5, 134.5,133.0, 127.6, 127.1, 113.9, 70.4, 45.6, 31.3, 30.1, 20.6, 15.9.

Example 4

Anti-alcohol 3d. Following the protocol used for preparation of 3a thedesired alcohol 3d was obtained as a colorless oil in 66% yield from 5g(S)-(+)-carvone. ¹H-NMR (270 MHz, CDCl₃) δ 7.65 (d, 1H), 7.31-7.24 (m,2H), 6.74-6.65 (m, 1H), 5.15-5.08 (dd, J=7.32, 4.75, 1H), 4.88 (s, 1H),3.26 (d, J=7.32 Hz, 1H), 3.01-2.95 (dd, J=4.75 Hz, 9.3 Hz, 1H),2.89-2.79 (m, 1H), 2.49 (m, 2H), 1.74 (s, 6H). ¹³C-NMR (270 MHz, CDCl₃)201.4, 144.8, 144.1, 139.1, 135.6, 133.6, 132.4, 130.4, 128.9, 127.2,114.1, 69.7, 54.2, 45.6, 30.2, 20.3, 15.9.

Example 5

Anti-alcohol 3e. Following the protocol used for preparation of 3a thedesired alcohol 3e was obtained in 65% yield (2.22 g, 7.25 mmol) from1.96 g (13.1 mmol) of (S)-(+)-carvone. ¹H NMR (300 MHz, CDCl₃) δ 8.27(d, J=7.6 Hz, 1H), 8.13 (d, J=7.5 Hz, OH), 7.94-7.78 (m, 3H), 7.62 (d,J=7.2 Hz, 1H), 7.60-7.41 (m, 5H), 6.72 (s, 1H), 5.58 (t, J=6.1 Hz, 1H),5.16 (s, 1H), 4.82 (d, J=18.7 Hz, 2H), 3.38 (d, J=6.5 Hz, 1H), 3.30-3.20(m, 1H), 2.72-2.61 (m, 1H), 2.61-2.32 (m, 2H), 1.84 (d, J=1.4 Hz, 3H),1.61 (s, 3H). ¹³C-NMR (75 MHz, CDCl₃) δ 201.86, 145.62, 143.77, 137.80,135.55, 131.34, 128.85, 128.75, 126.52, 126.05, 125.59, 125.50, 123.84,113.38, 71.11, 63.87, 44.66, 29.39, 21.09, 16.16. Calcd for C₂₁H₂₂O₂: C,82.32; H, 7.24. Found: C, 82.23; H, 7.36.

Example 6

Anti-alcohol 3f. Following the protocol used for preparation of 3a thedesired alcohol 3f was obtained in 28% yield (1.92 g, 7.5 mmol) from4.00 g (26.6 mmol) of (S)-(+)-carvone. ¹H NMR (300 MHz, CDCl₃) δ 8.50(d, J=4.8 Hz, 1H), 7.68 (td, J=1.7, 7.7 Hz, 1H), 7.51 (d, J=7.6 Hz, 1H),7.13 (dd, J=4.9, 7.4 Hz, 1H), 6.74-6.65 (m, 1H), 4.94 (dd, J=10.4, 11.8Hz, 2H), 4.84 (d, J=7.9 Hz, 1H), 3.90 (d, J=9.1 Hz, 1H), 3.50 (dd,J=2.1, 12.4 Hz, 1H), 3.13 (ddd, J=5.1, 10.5, 12.4 Hz, 1H), 2.65-2.47 (m,1H), 2.39 (dt, J=5.2, 9.5 Hz, 1H), 1.85 (s, 3H), 1.68 (s, 3H). ¹³C-NMR(75 MHz, CDCl₃) δ 200.91, 163.01, 148.18, 145.60, 144.46, 136.58,135.88, 121.65, 119.69, 114.48, 72.07, 54.52, 46.27, 31.26, 19.12,15.84. Calcd for C₁₆H₁₉NO₂: C, 74.68; H, 7.44. Found: C, 74.45; H, 7.50.

Example 7

Anti-alcohol 3g. Following the protocol used for preparation of 3a thedesired alcohol 3g was obtained as an oil in 47% yield (1.15 g, 4.7mmol) from 1.5 g (10.0 mmol) of (S)-(+)-carvone. ¹H-NMR (300 MHz, CDCl₃)δ 7.30 (dd, J=0.8, 1.8 Hz, 1H), 6.78 (ddd, J=1.3, 2.4, 6.1 Hz, 1H), 6.29(dd, J=1.8, 3.2 Hz, 1H), 6.16 (d, J=3.3 Hz, 1H), 5.20 (d, J=10.9 Hz,1H), 4.91-4.83 (m, 1H), 4.75 (dd, J=4.5, 11.0 Hz, 2H), 2.95 (dd, J=4.4,12.8 Hz, 1H), 2.64-2.50 (m, 1H), 2.50-2.34 (m, 1H), 2.33-2.19 (m, 1H),1.81 (dt, J=1.3, 2.4 Hz, 3H), 1.74 (s, 4H). ¹³C-NMR (75 MHz, CDCl₃) δ202.69, 154.94, 154.81, 144.03, 141.68, 135.78, 114.37, 110.23, 108.17,68.24, 52.10, 45.29, 31.17, 18.74, 15.74. Calcd for C₁₅H₁₈O₃: C, 73.15;H, 7.37. Found: C, 73.39; H, 7.43.

Example 8

Anti-alcohol 3h. Following the protocol used for preparation of 3a thedesired alcohol 3h was obtained in 50% yield (1.79 g, 6.52 mmol) from1.96 g (13.1 mmol) of (S)-(+)-carvone. ¹H-NMR (300 MHz, CDCl₃) δ 7.35(td, J=1.7, 7.6 Hz, 1H), 7.30-7.14 (m, 1H), 7.07 (dt, J=3.8, 7.5 Hz,1H), 6.96 (ddd, J=1.2, 8.2, 10.6 Hz, 1H), 6.72 (ddd, J=1.4, 3.6, 4.9 Hz,1H), 5.39 (dd, J=5.1, 8.1 Hz, 1H), 4.71 (dd, J=12.7, 14.1 Hz, 2H), 4.12(d, J=8.2 Hz, 1H), 3.03 (dd, J=5.1, 9.8 Hz, 1H), 2.84-2.68 (m, 1H),2.51-2.25 (m, 2H), 1.77 (s, 3H), 1.58 (s, 3H). ¹³C-NMR (75 MHz, CDCl₃) δ202.01, 161.92, 158.66, 145.11, 135.90, 129.28, 129.16, 129.11, 128.99,124.07, 115.49, 115.19, 113.26, 67.87, 53.89, 43.54, 30.52, 19.83,16.04. Calcd for C₁₇H₁₉FO₂: C, 74.43; H, 6.98. Found: C, 74.70; H, 7.03.

Example 9

Glycolate 4a. To a stirring solution of alcohol 3a (8.65 g, 25.80 mmol)in anhydrous dimethylformamide (DMF) (75 mL) was added 8.97 g (38.7mmol, 1.5 eq) Ag₂O, followed by dropwise addition of ethyl bromoacetate(4.29 mL, 38.7 mmol) at rt. After stirring ˜10 min, 2,6-lutidine (3.1mL, 38.7 mmol) was added very slowly (˜0.2 mL/h) via syringe pump andthe mixture was then allowed to stir at rt an additional 24 h. The crudesolution was then filtered through a short silica gel column and elutedwith diethyl ether. The filtrate was washed with 3N HCl (100 mL) andextracted with hexane (3×60 mL). The combined extracts were washed withsaturated aqueous NaHCO₃ and brine, then dried over MgSO₄, andconcentrated in vacuo. The crude mixture was purified via flashchromatography eluting with 20:80 EtOAc/hexanes to yield glycolate 4a asa yellow oil (8.47 g, 78%). ¹H-NMR (270 MHz, CDCl₃) δ 7.54-7.51 (m, 2H),7.37-7.35 (t, 1H), 7.18-7.16 (t, 1H), 6.61 (s, 1H), 5.23-5.21 (d, 1H),4.74-4.66 (d, 2H) 4.15-4.12 (m, 2H), 3.99-3.92 (d, 1H), 3.68-3.62 (d,1H), 3.00-2.97 (m, 2H), 2.53-2.52 (s, 1H), 2.38-2.31 (d, 1H), 1.82, (s,3H), 1.63 (s, 3H), 1.24-1.19 (t, 3H). ¹³C-NMR (270 MHz, CDCl₃) 198.3,169.9, 141.7, 129.8, 129.0, 128.8, 128.1, 127.6, 112.2, 80.7, 77.6,77.1, 76.6, 65.6, 64.9, 60.7, 55.4, 43.5, 28.4, 21.4, 16.3, 14.2.

Example 10

Glycolate 4b. Following the above procedure for preparation of 4a,glycolate 4b was obtained in 77% yield as a colorless oil from 1.61 g(4.80 mmol) alcohol 3b. ¹H-NMR (270 MHz, CDCl₃) δ 7.53-7.49 (d, 1H),7.45-7.42 (d, 1H), 7.33-7.27 (t, 1H), 7.18-7.07 (m, 1H), 6.64 (s, 1H),5.42-5.39 (d, 1H), 4.69-4.64 (d, 2H) 4.23-4.14 (m, 2H), 4.00-3.94 (d,J=16.43, 1H), 3.82-3.76 (d, J=16.43 Hz, 1H), 3.33-3.26 (m, 1H),2.97-2.92 (m, 2H), 2.43-2.36 (m, 1H), 1.72, (s, 3H), 1.50 (s, 3H),1.27-1.22 (t, 3H). ¹³C-NMR (270 MHz, CDCl₃) 197.8, 170.1, 146.1, 143.4,138.0, 135.5, 132.6, 129.6, 129.4, 127.6, 112.3, 79.4, 72.6, 68.0, 66.2,61.0, 55.7, 41.5, 28.2, 21.2, 16.3, 14.3.

Example 11

Glycolate 4c. Following the above procedure, glycolate 4c was obtainedin 83% yield as a colorless oil from 7.06 g (21.71 mmol) alcohol 3c.¹H-NMR (270 MHz, CDCl₃) δ 7.53-7.49 (d, 1H), 7.44-7.40 (d, 1H),7.30-7.26 (m, 1H), 6.64 (s, 1H), 5.20-5.18 (d, 1H), 4.81-4.76 (d, 2H)4.20-4.11 (m, 2H), 4.03-3.97 (d, J=16.03 Hz, 1H), 3.72-3.66 (d, J=16.03Hz, 1H), 3.31-3.25 (m, 1H), 2.94-2.90 (m, 2H), 2.44-2.34 (m, 1H), 1.72,(s, 3H), 1.48 (s, 3H), 1.31-1.26 (t, 3H). ¹³C-NMR (300 MHz, CDCl₃)198.9, 170.0, 143.5, 129.8, 129.5, 127.4, 127.0, 112.5, 70.7, 68.1,61.0, 55.1, 41.6, 28.5, 16.2, 14.2.

Example 12

Glycolate 4d. Following the above procedure, glycolate 4d was obtainedin 80% yield as a colorless oil from 9.48 g (29.15 mmol) alcohol 3d.¹H-NMR (270 MHz, CDCl₃) δ 7.53-7.49 (d, 1H), 7.44-7.40 (d, 1H),7.30-7.26 (m, 1H), 6.64 (s, 1H), 5.20-5.18 (d, 1H), 4.81-4.76 (d, 2H)4.20-4.11 (m, 2H), 4.03-3.97 (d, J=16.03 Hz, 1H), 3.72-3.66 (d, J=16.03Hz, 1H), 3.31-3.25 (m, 1H), 2.94-2.90 (m, 2H), 2.44-2.34 (m, 1H), 1.72,(s, 3H), 1.48 (s, 3H), 1.31-1.26 (t, 3H). ¹³C-NMR (300 MHz, CDCl₃)198.9, 170.0, 143.5, 129.8, 129.5, 127.4, 127.0, 112.5, 70.7, 68.1,61.0, 55.1, 41.6, 28.5, 16.2, 14.2.

Example 13

Glycolate 4e. Following the above procedure, glycolate 4e was obtainedas an oil in 25% yield (8.17 g, 25.13 mmol) from alcohol 3e (3.27 mmol).¹H-NMR (300 MHz, CDCl₃) δ 8.09 (d, J=7.6 Hz, 1H), 7.91-7.83 (m, 1H),7.79 (d, J=8.2 Hz, 1H), 7.60 (d, J=7.0 Hz, 1H), 7.56-7.41 (m, 3H), 6.75(s, 1H), 5.86 (d, J=5.1 Hz, 1H), 4.59 (s, 2H), 4.13 (d, J=16.4 Hz, 1H),3.92 (d, J=10.3 Hz, 1H), 3.70 (s, 3H), 3.28 (dd, J=4.4, 9.8 Hz, 1H),3.09-2.90 (m, 2H), 2.47-2.31 (m, 1H), 1.72 (s, 3H), 1.27 (s, 3H).¹³C-NMR (75 MHz, CDCl₃) δ 198.92, 170.89, 146.63, 144.20, 135.41,134.25, 133.96, 131.11, 129.22, 128.72, 126.54, 125.73, 125.26, 125.22,122.98, 112.11, 79.47, 66.40, 55.66, 51.85, 41.25, 29.00, 20.97, 16.37.HRMS Calcd for C₂₄H₂₆O₄Na: 401.1729. Found: 401.1719.

Example 14

Glycolate 4f. Following the above procedure, glycolate 4f was obtainedas an oil in 86% yield (1.43 g, 4.34 mmol) from 1.3 g (5.05 mmol) ofalcohol 3f. ¹H-NMR (300 MHz, CDCl₃) δ 8.50 (d, J=4.8, 1H), 7.70 (dt,J=7.0, 20.6 Hz, 2H), 7.23-7.10 (m, 1H), 6.63 (s, 1H), 4.83 (s, 1H), 4.75(d, J=5.2 Hz, 1H), 4.20 (d, J=16.2 Hz, 1H), 3.93 (d, J=16.2 Hz, 1H),3.69 (s, 3H), 3.17 (dd, J=5.2, 8.0 Hz, 1H), 2.89-2.70 (m, 1H), 2.50 (m,2H), 1.80-1.71 (m, 6H).

¹³C-NMR (75 MHz, CDCl₃) δ 197.99, 170.30, 160.59, 148.74 145.81, 142.32,136.52, 135.60, 122.39, 121.32, 113.28, 83.42, 67.59, 55.15, 51.80,44.20, 29.28, 20.03, 16.15. Calcd for C₁₉H₂₃NO₄: C, 69.28; H, 7.04.Found: C, 68.99; H, 7.09.

Example 15

Glycolate 4g. Following the above procedure, glycolate 4g was obtainedas an oil in 48% yield (679 mg, 2.13 mmol) from 1.1 g (4.46 mmol) ofalcohol 3g. ¹H-NMR (300 MHz, CDCl₃) δ 7.40 (dd, J=0.7 Hz, 1.7, 1H),6.68-6.59 (m, 1H), 6.33-6.20 (m, 2H), 4.88 (d, J=7.6 Hz, 1H), 4.74 (d,J=1.2 Hz, 1H), 4.66 (s, 1H), 4.12 (d, J=16.8 Hz, 1H), 3.96-3.87 (m, 1H),3.73-3.66 (m, 3H), 3.21 (dd, J=4.4, 9.4 Hz, 1H), 3.09 (dd, J=4.4, 7.5Hz, 1H), 2.98-2.81 (m, 1H), 2.42 (ddd, J=2.5, 5.6, 19.6 Hz, 1H), 1.68(s, 7H). ¹³C-NMR (75 MHz, CDCl₃) δ 198.03, 171.00, 151.32, 145.97,143.28, 143.06, 112.17, 110.34, 110.18, 74.74, 65.50, 54.79, 51.92,41.73, 28.02, 21.52, 16.28. Calcd. for C₁₈H₂₂O₅: C, 67.91; H, 6.97.Found: C, 67.71; H, 6.96.

Example 16

Glycolate 4h. Following the above procedure, glycolate 4h was obtainedas an oil in 19% yield (0.25 g, 0.71 mmol) from 1.05 g of alcohol 3h(3.8 mmol). ¹H NMR (300 MHz, CDCl₃) δ 7.44 (td, J=1.8, 7.5 Hz, 1H),7.31-7.21 (m, 2H), 7.15 (t, J=7.0 Hz, 1H), 6.97 (dd, J=8.4, 10.1 Hz,1H), 6.66 (s, 1H), 5.26 (d, J=7.6 Hz, 1H), 4.68 (d, J=11.6 Hz, 2H), 4.08(d, J=16.5 Hz, 1H), 3.87 (d, J=16.5 Hz, 1H), 3.73 (s, 3H), 3.31 (s, 1H),3.06-2.80 (m, 2H), 2.43 (d, J=19.6 Hz, 1H), 1.72 (s, 3H), 1.59 (s, 3H).¹³C-NMR (75 MHz, CDCl₃) δ 198.18, 170.72, 162.47, 159.21, 146.07,143.16, 135.39, 129.84, 128.75, 126.14, 125.97, 124.48, 124.43, 115.46,115.18, 112.2574.74, 66.05, 56.64, 51.90, 41.50, 27.91, 21.48, 16.24.Calcd for C₂₀H₂₃FO₄: C, 69.35; H, 6.69. Found: C, 68.98, H, 6.61.

Example 17

Isobenzofuran 5a. To a 0.1 M solution of glycolate 4a (4.13 g, 9.8 mmol)in dry

THF (100 mL) was added potassium bis(trimethylsilylamide) (KHMDS) (23.53mL, 11.76 mmol, 1.1 eq, 0.5 M soln. in toluene) quickly at −78° C.,followed immediately by rapid addition of 1.2 eq HOAc (0.673 mL, 11.76mmol). The reaction mixture was allowed to warm to rt before water (100mL) was added and the organic layer was extracted with ether (3×60 mL).The combined extracts were washed with brine and dried over MgSO₄,filtered and concentrated in vacuo. The crude product was purified viaflash chromatography with 10:90 EtOAc/hexanes to yield the desiredcycloaldol product 5a as a colorless oil (3.30 g, 7.84 mmol, 80%).¹H-NMR (270 MHz, CDCl₃) δ 8.26-8.22 (d, 1H), 7.50-7.48 (d, 1H),7.40-7.38 (t, 1H), 7.15-7.13 (t, 1H), 5.73 (s, 1H), 5.22-5.18 (d, 1H),4.80 (s, 2H), 4.42 (s, 1H), 4.28-4.25 (m, 2H), 2.74-2.69 (m, 1H),2.58-2.42 (d, 1H), 2.28-2.27 (m, 2H), 1.82 (s, 3H), 1.53 (s, 3H),1.33-1.28 (t, 3H). ¹³C-NMR (270 MHz, CDCl₃) 171.0, 147.0, 139.4, 132.9,132.4, 130.4, 129.7, 128.3, 125.6, 123.9, 112.2, 82.6, 81.9, 80.7, 61.3,55.5, 39.1, 27.7, 21.4, 17.7, 14.2.

Example 18

Isobenzofuran 5b. Following the same procedure used for preparing 5a,isobenzofuran 5b was obtained in 73% yield as a pale yellow oil from0.77 g (1.83 mmol) glycolate 4b. ¹H-NMR (270 MHz, CDCl₃) δ 8.31-8.26 (d,1H), 7.54-7.49 (d, 1H), 7.46-7.39 (t, 1H), 7.20-7.13 (t, 1H), 5.79-5.74(m, 1H), 5.27-5.21 (d, 1H), 4.83 (s, 2H), 4.45 (s, 1H), 4.35-4.23 (m,2H), 3.09 (s, 1H), 2.79-2.71 (dd, 1H), 2.60-2.47 (m, 1H), 2.35-2.22 (m,2H), 1.86 (s, 3H), 1.57 (s, 3H), 1.39-1.30 (t, 3H). ¹³C-NMR (270 MHz,CDCl₃) 171.0, 146.9, 139.4, 132.9, 132.4, 130.4, 129.7, 128.3, 125.6,123.8, 112.1, 82.6, 81.8, 80.6, 61.3, 55.5, 39.0, 27.7, 21.4, 17.7,14.15.

Example 19

Isobenzofuran 5c. Following the same procedure used for preparing 5a,isobenzofuran 5c was obtained in 77% yield as a pale yellow oil from3.39 g (8.24 mmol) glycolate 4c. ¹H-NMR (270 MHz, CDCl₃) δ 8.26-8.16 (d,1H), 7.42-7.34 (d, 1H), 7.32-7.21 (t, 1H), 5.71 (s, 1H), 5.34-5.26 (d,1H), 4.79 (s, 2H), 4.42 (s, 1H), 4.34-4.14 (m, 2H), 3.06 (s, 1H),2.72-2.61 (dd, 1H), 2.49-2.16 (m, 3H), 1.81 (s, 3H), 1.54 (s, 3H),1.35-1.20 (t, 3H). ¹³C-NMR (270 MHz, CDCl₃) 171.1, 146.7, 140.8, 132.9,132.6, 131.5, 130.1, 128.4, 128.1, 125.7, 112.5, 81.9, 80.9, 80.7, 61.4,55.4, 39.7, 27.7, 21.3, 17.7, 14.2.

Example 20

Isobenzofuran 5d. Following the same procedure used for preparing 5a,isobenzofuran 5d was obtained in 74% yield as a pale yellow oil from4.73 g (11.50 mmol) glycolate 4d. ¹H-NMR (270 MHz, CDCl₃) δ 8.30-8.25(d, 1H), 7.39-7.32 (m, 2H), 5.75 (S, 1H), 5.29-5.23 (d, 2H), 4.84 (s,2H), 4.46 (s, 1H), 4.37-4.22 (m, 2H), 3.16 (s, 1H), 2.73-2.64 (m, 1H),2.50-2.19 (m 3H), 1.85 (s, 3H), 1.59 (s, 3H), 1.38-1.31 (t, 2H).

Example 21

Isobenzofuran 5e. Following the same procedure used for preparing 5a,isobenzofuran 5e was obtained in 59% yield (0.22 g, 0.59 mmol) from 0.41g (1.08 mmol) of glycolate 4e. ¹H-NMR (300 MHz, CDCl₃) δ 8.34 (d, J=8.0Hz, 1H), 8.16 (d, J=7.2 Hz, 1H), 7.86 (t, J=9.1 Hz, 2H), 7.52 (dt,J=6.9, 14.5 Hz, 3H), 5.76 (s, 1H), 5.62 (d, J=7.2 Hz, 1H), 4.91 (s, 1H),4.83 (s, 1H), 4.59 (s, 1H), 3.85 (s, 3H), 3.26 (s, 1H), 3.03 (t, J=7.2Hz, 1H), 2.39 (dd, J=6.5, 12.7 Hz, 1H), 2.29 (s, 2H), 1.87 (s, 3H), 1.55(s, 3H). ¹³C-NMR (75 MHz, CDCl₃) δ 172.29, 146.85, 135.72, 133.92,133.46, 131.85, 128.99, 128.86, 126.11, 125.74, 125.63, 125.60, 125.52,123.60, 113.03, 82.77, 81.51, 80.99, 53.68, 52.36, 42.04, 28.22, 20.68,17.79. Calcd for C₂₄H₂₆O₄: C, 76.17; H, 6.92. Found: C, 76.09; H, 6.95.

Example 22

Isobenzofuran 5f. Following the same procedure used for preparing 5a,isobenzofuran 5f was obtained in 81% yield (0.30 g, 0.90 mmol) from 0.36g (1.10 mmol) of glycolate 4f. ¹H-NMR (300 MHz, CDCl₃) δ 8.53 (d, J=4.0Hz, 1H), 7.64 (td, J=1.8, 7.7 Hz, 1H), 7.19 (ddd, J=3.6, 6.0, 7.8 Hz,2H), 5.58 (d, J=1.5 Hz, 1H), 4.88 (s, 1H), 4.82 (s, 1H), 4.55 (s, 1H),3.69 (s, 3H), 2.55-2.32 (m, 2H), 2.07 (d, J=3.1, 2H), 1.89-1.73 (m, 3H),1.50 (s, 3H). ¹³C-NMR (75 MHz, CDCl₃) δ 170.36, 159.63, 149.19, 146.18,137.80, 133.65, 125.22, 123.42, 123.23, 113.50, 87.12, 84.83, 82.53,54.58, 52.07, 46.56, 30.98, 19.36, 17.95. HRMS Calcd for C_(I9)H₂₃NO₄:C, 69.28; H, 7.04. Found: C, 68.98; H, 6.86.

Example 23

Enone Ester 6a. To a solution of isobenzofuran 5a (0.37 g, 0.87 mmol) inanhydrous CH₂Cl₂ (9.0 mL) was added premixed mixture of pyridiniumchlorochromate (PCC) (2.0 g, 8.7 mmol)/silica gel (2.0 g) and 3-4 dropsof anhydrous toluene. The reaction mixture was allowed to stir at rt for24 h, then was filtered through a short silica gel column with ether(100 mL). The eluant was concentrated in vacuo and the dark brownresidue was purified via flash chromatography over silica gel (85/15,hexane/EtOAc) to afford enone 6a as a white solid (0.2 g, 55%). ¹H-NMR(270 MHz, CDCl₃) δ 8.12-7.90 (d, 1H), 7.50-7.30 (m, 2H), 7.12-7.08 (d,1H), 5.30 (s, 1H), 5.20-5.10 (d, 1H), 4.70 (s, 1H), 4.40 (s, 1H),4.28-4.18 (m, 2H), 3.30-3.10 (m, 1H), 2.85-2.70 (m, 1H), 2.45-2.31 (m,2H), 1.90 (s, 3H), 1.40 (t, 3H), 1.20 (s, 3H). ¹³C-NMR (270 MHz, CDCl₃)198.0, 171.0, 146.0, 142.0, 138.0, 132.9, 130.4, 130.0, 129.7, 128.3,125.6, 114.2, 85.2, 78.5, 62.3, 51.5, 47.5, 42.0, 18.5, 14.7, 12.2.

Example 24

Enone ester 6b. Following the same procedure used for preparing 6a,enone 6b was obtained in 54% yield as a white solid from 0.124 g (0.294mmol) isobenzofuran 5b. ¹H-NMR (270 MHz, CDCl₃) δ 8.04-7.99 (d, 1H),7.51-7.47 (d, 1H), 7.45-7.38 (t, 1H), 7.21-7.15 (t, 1H), 5.30-5.27 (m,1H), 5.24-5.19, (d, 1H), 4.74 (s, 1H), 4.47 (s, 1H), 4.36-4.27 (q,J=7.11 Hz, 2H), 3.31-3.21 (m, 1H), 2.88-2.77 (m, 1H), 2.49-2.34 (m, 2H),1.93 (s, 1H), 1.41-1.34 (t, J=7.11 Hz, 3H), 1.27 (s, 3H). ¹³C-NMR (270MHz, CDCl₃) 183.6, 156.0, 142.3, 132.6, 129.9, 127.9, 113.9, 85.0, 78.1,66.6, 62.0, 58.0, 46.7, 42.4, 22.7, 18.02, 14.2, 11.4.

Example 25

Enone ester 6c. Following the same procedure used for preparing 6a,enone 6c was obtained in 56% yield as a white solid from 0.0.4 g (0.972mmol) isobenzofuran 5c. ¹H-NMR (270 MHz, CDCl₃) δ 7.94-7.91 (d, 1H),7.36-7.35 (d, 1H), 7.28-7.24 (t, 1H), 5.28-5.23 (d, 2H), 4.61 (s, 1H),4.41 (s, 1H), 4.26-4.18 (m, 2H), 3.21-3.08 (m, 1H), 2.80-2.65 (m, 1H),2.35-2.23 (m, 2H), 1.85 (s, 3H), 1.25 (t, 3H), 1.20 (s, 3H). ¹³C-NMR(270 MHz, CDCl₃) 197.5, 169.8, 154.7, 142.1, 139.0, 132.5, 129.9, 127.4,113.4, 82.6, 62.0, 50.4, 46.9, 42.3, 18.2, 14.1, 11.1.

Example 26

Enone ester 6d. Following the same procedure used for preparing 6a,enone 6d was obtained in 67% yield as a white solid from 0.17 g (0.41mmol) isobenzofuran 5d. ¹H-NMR (270 MHz, CDCl₃) δ 7.97-7.94 (d, 1H),7.36-7.23 (m, 2H), 5.23 (s, 1H), 5.15-5.11 (d, 1H), 4.66 (s, 1H), 4.46(s, 1H), 4.31-4.21 (m, 2H), 3.21-3.11 (m, 1H), 2.80-2.69 (m, 1H),2.46-2.29 (m, 2H), 1.88 (s, 3H), 1.35-29 (t, 3H). 1.26 (s, 3H). ¹³C-NMR(270 MHz, CDCl₃) 198.1, 169.8, 155.5, 142.4, 135.6, 134.7, 130.5, 130.4,129.0, 127.8, 114.0, 82.0, 62.2, 50.2, 46.7, 42.3, 18.2, 14.3, 11.5.

Example 27

Tosylhydrazone 7. HOAc (0.01 g, 0.16 mmol) was added to a solution ofenone ester 6a (0.10 g, 0.24 mmol) and tosylhydrazide (0.06 g, 0.31mmol) in CH₂Cl₂ (2.20 mL). The reaction mixture stirred at rt for 24 h,then was washed with water (5.00 mL), dried over MgSO₄, filtered andconcentrated in vacuo. The residue was purified via flash chromatographyon silica gel(80:20, Hexane/EtOAc) to afford tosylhydrazone 7 as whitesolid in 92% yield (0.13 g, 0.22 mmol); mp 162-165° C. ¹H-NMR (400 MHz,CDCl₃) δ 8.02-7.79 (m, 3H), 7.46 (d, J=9.5 Hz, 2H), 7.36 (t, J=8.7 Hz,3H), 7.14 (t, J=7.7 Hz, 1H), 5.28 (s, 1H), 5.07 (d, J=9.8 Hz, 1H), 4.70(s, 1H), 4.48 (s, 1H), 4.25 (q, J=7.1 Hz, 2H), 2.99 (s, 1H), 2.58-2.34(m, 6H), 2.10-1.85 (m, 4H), 1.34 (t, J=7.1 Hz, 3H), 1.22 (s, 3H).¹³C-NMR (101 MHz, CDCl₃) δ 170.44, 153.31, 144.28, 143.92, 142.66,138.39, 135.01, 132.51, 129.73, 129.69, 129.49, 128.25, 127.80, 127.78,124.74, 114.17, 85.49, 78.23, 77.35, 77.04, 76.72, 61.70, 49.10, 44.80,29.37, 21.66, 18.00, 14.18, 13.07. IR (film) 3210, 3070, 2980, 2919,2256, 1739, 1442, 1402 cm*HRMS Calcd for C₂₈H₃₁BrN₂NaO₅S⁺: 587.1215.Found: 587.1201.

Example 28

Ester 8. Catecholborane (0.10 mL, 0.80 mmol)) was added to a solution oftosylhydrazone 7 (0.40 g, 0.70 mmol) in CHCl₃ (3.00 mL) at 0° C. Thereaction mixture was stirred at 0° C. for 1 h, then NaOAc.3H₂O (0.19 g,1.30 mmol) was added in one portion. The reaction mixture was maintainedfor 1 h at 0° C., diluted with CHCl₃ (1.80 mL), and heated under refluxfor 12 h. The mixture was then cooled to rt and filtered through a padof Celite. The filtrate was concentrated in vacuo and the residue waspurified via flash chromatography over silica gel (90:10, hexanes/EtOAc)to afford reduced ester 8 as white solid in 68% yield (0.17 g, 0.48mmol); mp 37-40° C.; ¹H-NMR (270 MHz, CDCl₃) δ 7.97 (d, J=6.2 Hz, 1H),7.49 (d, J=8.0 Hz, 1H), 7.36 (t, J=7.0 Hz, 1H), 7.12 (t, J=7.6 Hz, 1H),5.59 (s, 1H), 5.34 (d, J=6.4 Hz, 1H), 4.79 (s, 2H), 4.38 (d, J=6.4 Hz,1H), 4.25 (dd, J=4.9, 7.1 Hz, 2H), 3.01 (s, 1H), 2.37 (m, 3H), 2.21-2.04(m, 1H), 1.73 (s, 3H), 1.60 (s, 4H), 1.31 (t, J=7.1 Hz, 3H). ¹³C-NMR (67MHz, CDCl₃) δ 173.01, 146.94, 140.85, 132.70, 130.21, 129.58, 129.37,128.04, 123.25, 123.10, 112.16, 83:50, 81.30, 77.56, 77.09, 76.62,61.33, 49.14, 47.31, 39.39, 28.04, 22.08, 21.00, 14.22.

Example 29

Methyl ester 5a.i. Into a 25 mL round bottom flask was placed 100 mg(0.24 mmol) ethyl ester 5a, 15 mL methanol (11.85 g, 370 mmol) and amagnetic stir bar. To the mixture was added 0.04 g (0.28 mmol, 1.2 eq)K₂CO₃ and the reaction was allowed to stir at room temperature. After˜20 min the mixture turned from a very light yellow color to dark orangeand TLC revealed no starting material remained. The mixture was thentransferred to a seperatory funnel and diluted with 10 mL H₂O andextracted with CH₂Cl₂ (3×20 mL). The combined organic extracts werewashed with brine (50 mL), extracted with CH₂Cl₂ (40 mL), dried overMgSO₄, and concentrated in vacuo. The crude product was filtered througha short silica gel column eluted with EtOAc/Hexanes (50:50) to delivermethyl ester 5a.i as a colorless oil (94.8 mg, 23.3 mg, 97%). ¹H-NMR(270 MHz, CDCl₃) δ8.32-8.28 (d, 1H), 7.55-7.52 (d, 1H), 7.48-7.42 (t,1H), 7.22-7.16 (t, 1H), 5.82-5.78 (m, 1H), 5.27-5.23 (d, 1H), 4.87-4.84(m, 2H), 4.51 (s, 1H), 3.85 (s, 3H), 2.93 (s, 1H), 2.80-2.73 (m, 1H),2.65-2.53 (m, 1H), 2.38-2.26 (m, 2H), 1.88 (s, 1H), 1.59 (s, 1H).

Example 30

Cyclopropylmethyl ester 5a.ii. Into an 8 mL microwave vessel was placed0.14 g (0.33 mmol) ethyl ester 5a and a magnetic stir bar. Added to thestarting material was 2.66 mL (100 eq, 33.22 mmol) cyclopropyl methanolfollowed by 0.063 g (0.75 eq, 0.241) Bu₂SnO. The vessel was sealed andreacted in a microwave reactor for 30 min. at 150° C. and 300 watts ofpower with continuous stirring. The reaction was diluted with 10 mLethyl acetate and transferred to a separatory funnel, washed withsaturated NaHCO₃ (30 mL) and extracted with EtOAc (3×20 mL). Thecombined organic extracts were then washed with brine, dried over MgSO₄,and concentrated in vacuo. The crude product was purified on a silicagel column eluted with EtOAc/Hexanes (10:90) to deliver ester 5a.ii as acolorless oil (0.125 g, 0.28 mmol, 84%). ¹H-NMR (270 MHz, CDCl₃)δ8.31-8.26 (d, 1H), 7.55-7.50 (d, 1H), 7.46-7.39 (m, 1H), 7.21-7.14 (m,1H), 5.81-5.75 (m, 1H), 5.28-5.23 (d, 1H), 4.84 (s, 2H), 4.48 (s, 1H),4.15-4.01 (m, 2H), 3.16 (s, 1H), 2.80-2.74 (m, 1H), 2.59-2.45 (m, 1H),2.36-2.24 (m, 2H), 1.87 (s, 3H), 1.57 (s, 3H), 1.29-1.14 (m, 1H),0.65-0.57 (m, 2H), 0.38-0.31 (m, 2H). ¹³C-NMR (270 MHz, CDCl₃) 171.0,146.8, 139.4, 132.9, 132.4, 130.4, 129.7, 128.3, 125.7, 112.2, 82.7,81.9, 80.7, 70.2, 55.5, 39.5, 27.8, 21.27, 17.7, 9.6, 3.4.

Example 31

Cyclopentylmethyl ester 5a.iii. Ester 5a.iii was prepared as above for5a.ii in 91% yield (0.11 g, 0.24 mmol) from 0.11 g (0.26 mmol) of ester5a. ¹H-NMR (270 MHz, CDCl₃) 8.29 (d, J=6.2 Hz, 1H), 7.53 (d, J=8.0 Hz,1H), 7.44 (t, J=7.5 Hz, 1H), 7.19 (dd, J=4.5, 10.8 Hz, 1H) 5.79 (s, 1H),5.25 (d, J=9.0 Hz, 1H), 4.85 (s, 2H), 4.49 (s, 1H), 4.17-4.04 (m, 2H),2.99 (s, 1H), 2.76 (dd, J=5.0, 9.1 Hz, 1H), 2.64-2.52 (m, 1H), 2.32 (d,J=7.3 Hz, 3H); 1.89-1.74 (m, 6H), 1.68-1.54 (m, 8H), 1.39-1.26 (m, 3H).¹³C-NMR (67 MHz, CDCl₃) 171.1, 147.1, 139.5, 132.9, 132.4, 130.3, 129.7,128.3, 125.5, 123.8, 112.0, 82.5, 81.9, 80.6, 69.3, 55.5, 38.5, 38.3,29.3, 27.5, 25.4, 21.5, 17.6.

Example 32

Acid 9. To a solution of 3:2:1 THF/H₂O/MeOH and 0.180 g (0.43 mmol)ester 5a was added 0.02 g (0.85 mmol, 2 eq) LiOH at rt. The reactionmixture was allowed to stir at rt ˜1.5 h until no starting material wasobserved by TLC. The solution was then titrated to pH˜2 with 3N HCl anddiluted with CH₂Cl₂ (30 mL). The mixture was transferred to a separatoryfunnel, extracted with CH₂Cl₂ (3×30 mL), and the combined extractswashed with brine (40 mL). The combined organic extracts were dried overMgSO₄ and concentrated in vacuo to yield a yellow foam which wasfiltered through a plug of silica gel (50:50 EtOAc/Hexane) to produceacid 9 as a white powder (159 mg, 0.406 mmol 95%). ¹H-NMR (270 MHz,CDCl₃) δ 8.14-8.09 (d, 1H), 7.59-7.54 (d, 1H), 7.47-7.40 (t, 1H),7.25-7.18 (t, 1H), 5.81 (s, 1H), 5.28 (s, 1H), 5.25 (d, 1H), 4.85 (s,2H), 4.51 (s, 1H), 2.79-2.72 (dd, 1H), 2.60-2.47 (m, 1H), 2.35-2.24 (m,2H), 1.94 (s, 3H), 1.59 (s, 3H). ¹³C-NMR (270 MHz, CDCl₃) 146.6, 138.6,132.9, 132.7, 130.0, 129, 128.4, 125.9, 124.1, 112.4, 82.7, 55.6, 27.6,21.4, 17.7.

Example 33

Diene 10a. Preparation was carried out as above for 5a from glycolate 4a(3.5 g, 8.31 mmol), but the reaction mixture was allowed to stir for 5minutes before quenching with HOAc. Purification of the crude productvia flash chromatography over silica gel (10:90 EtOAc/hexanes) delivereddiene 10a as a yellow oil in 15% yield (0.42 g, 1.27 mmol) as well asisobenzofuran 5a in 41% yield (1.43 g, 3.41 mmol). ¹H-NMR (400 MHz,CDCl₃) δ 7.64 (d, J=6.5, 1H), 7.34-7.08 (m, 3H), 6.62 (s, 1H), 4.95 (s,1H), 4.73 (s, 1H), 3.62 (s, 1H), 2.74-2.48 (m, 1H), 2.19 (s, 1H),1.95-1.82 (m, 2H), 1.77 (s, 2H), 1.59 (s, 1H). ¹³C-NMR (101 MHz, CDCl₃)δ 189.40, 145.37, 141.23, 139.35, 136.66, 136.31, 134.83, 132.82,129.65, 129.62, 127.04, 125.00, 114.21, 77.35, 77.03, 76.71, 43.88,29.72, 29.18, 21.78, 16.34.

Example 34

Diene 10e. Diene 10e was prepared as above for 10a and was obtained in33% yield (0.10 g, 0.33 mmol) from 0.41 g (1.08 mmol) of glycolate 4e.¹H-NMR (300 MHz, CDCl₃) δ 8.34 (d, J=8.0 Hz, 1H), 8.16 (d, J=7.2 Hz,1H), 7.86 (dd, J=8.2, 10.1 Hz, 2H), 7.61-7.42 (m, 3H), 5.76 (s, 1H),5.62 (d, J=7.2 Hz, 1H), 4.87 (d, J=26.1 Hz, 2H), 4.59 (s, 1H), 3.83 (s,3H), 3.26 (s, 1H), 3.03 (t, J=7.2 Hz, 1H), 2.39 (dd, J=6.5, 12.7 Hz,1H), 2.34-2.21 (m, 2H), 1.87 (s, 3H), 1.55 (s, 3H). ¹³C-NMR (75 MHz,CDCl₃) δ 189.94, 146.02, 141.49, 140.18, 136.87, 133.88, 133.65, 133.24,132.32, 128.97, 128.70, 126.58, 126.32, 126.02, 125.37, 124.92, 114.15,44.28, 29.57, 22.06, 16.59. Calcd for C₂₂H₂₂O: C, 87.38; H, 7.33. Found:C, 87.78; H, 7.26.

Example 35

Diene 10g. Diene 10g was prepared as above for 10a and was obtained in41% yield (0.05 g, 0.19 mmol) from 0.15 g (0.46 mmol) of glycolate 4g.¹H-NMR (300 MHz, CDCl₃) δ 7.50 (d, J=1.6 Hz, 1H), 7.42 (s, 1H),6.70-6.61 (m, 1H), 6.58 (d, J=3.4 Hz, 1H), 6.47 (dd, J=1.8, 3.4 Hz, 1H),4.87-4.74 (m, 1H), 4.65 (s, 1H), 4.37 (d, J=6.1 Hz, 1H), 2.77-2.45 (m,2H), 1.86 (s, 3H), 1.81 (s, 3H). ¹³C-NMR (75 MHz, CDCl₃) δ 188.77,152.31, 145.57, 144.27, 141.82, 136.68, 134.43, 122.87, 116.05, 112.65,112.24, 43.29, 28.94, 22.11, 16.77. Calcd for C₁₆H₁₈O₂: C, 79.31; H,7.49. Found: C, 79.23; H, 7.24.

Example 36

Diene 10h. Diene 10h was prepared as above for 10a and was obtained in45% yield (0.07 g, 0.26 mmol) from 0.20 g (0.58 mmol) of glycolate 4h.¹H NMR (300 MHz, CDCl₃) δ 7.68 (s, 1H), 7.33 (dt, J=4.5, 14.2 Hz, 2H),7.12 (dd, J=8.3, 16.2 Hz, 2H), 6.67-6.59 (m, 1H), 4.96 (s, 1H), 4.74 (s,1H), 3.75 (s, 1H), 2.72-2.50 (m, 2H), 1.90 (s, 3H), 1.81 (s, 3H).¹³C-NMR (75 MHz, CDCl₃) δ 189.45, 162.81, 159.50, 145.36, 141.33,140.24, 136.68, 130.42, 130.31, 129.93, 129.90, 128.54, 128.49, 124.03,123.98, 123.86, 115.88, 115.59, 114.30, 44.28, 29.24, 21.92, 16.50.Calcd for C₁₈H₁₉FO: C, 79.97; H, 7.08. Found: C, 79.58; H, 6.72.

Example 37

Isobenzofuran 51. A solution of isobenzofuran 5f (0.13 g, 0.394 mmol)and iodomethane (ca. 20 eq) was heated under reflux in MeCN. After 3days, the reaction mixture was cooled and concentrated under the reducedpressure to give pyridinium iodide 51 in 98% yield (0.18 g, 0.39 mmol).¹H-NMR (300 MHz, CDCl₃) δ 9.48 (d, J=8.4 Hz, 1H), 8.82 (d, J=6.2 Hz,1H), 8.33 (t, J=8.0 Hz, 1H), 7.85 (t, J=6.2 Hz, 1H), 5.61 (s, 1H), 5.48(s, 1H), 5.05 (d, J=7.6 Hz, 2H), 4.71 (d, J=1.8 Hz, 1H), 4.38 (s, 3H),3.94 (d, J=12.1 Hz, 1H), 3.85-3.69 (m, 4H), 2.63 (dd, J=5.6, 11.8 Hz,1H), 2.46 (d, J=17.9 Hz, 1H), 1.80 (d, J=16.3 Hz, 6H). ¹³C-NMR (75 MHz,CDCl₃) δ 170.15, 159.52, 145.70, 144.41, 133.23, 130.75, 126.42, 125.31,116.53, 86.99, 83.31, 80.34, 52.75, 52.38, 46.23, 45.65, 29.89, 19.97,18.17. Calcd for C₂₀H₂₆INO₄: C, 50.97; H, 5.56. Found: C, 51.11; H,5.65.

Example 38

Cell Proliferation Studies

Inhibition of cell proliferation was assessed by the MTT assay. Thisassay is based on the ability of a mitochondrial dehydrogenase enzymefrom viable cells to cleave the tetrazolium rings of the pale yellow3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) andform a dark blue formazan crystal product which is largely impermeableto cell membranes, thus resulting in its accumulation within healthycells.(22) Solubilization of the cells by the addition of solventresults in the liberation of the crystals which are solubilized. Thelevel of the formazan product created is directly proportional to thenumber of living cells.(23,24)

All stock solutions of compounds were made at 10 mM in dimethylsulfoxide (DMSO). DMSO and MTT were from Sigma Chemical Co. (St. Louis,Mo.). Cell culture reagents were obtained from Life Technologies(Carlsbad, Calif.). The KB-3 human carcinoma cell line was maintained inmonolayer culture at 37° C. and 5% CO₂ in Dulbecco's Modified Eagle'sMedium, supplemented with 10% fetal bovine serum, 2 mM L-glutamine, 50units/mL penicillin, and 50 μg/mL streptomycin.

KB-3 cells (2000/well) were plated in 96-well dishes, and after 24 hwere treated with 0.1 nM-100 μM of the compound in question. The finalconcentration of DMSO did not exceed 1%, and controls received vehiclealone. After 96 h, cells were incubated with 50 μg MTT/well/0.2 mL for 4h at 37° C., the media was removed, and the formazan crystals weredissolved in 0.15 mL of DMSO. Absorbance at 570 nm was measured with anELx800™ Microplate Reader (Bio Tek Instruments, Inc., Winooski, Vt.).All treatments were performed in triplicate. Results are shown in FIGS.1A-D as a percentage of control samples (mean±SD). IC₅₀ is theconcentration that reduced survival to 50% of the control (no drug).

The present invention has been described hereinabove with reference toparticular examples for purposes of clarity and understanding ratherthan by way of limitation. It should be appreciated that certainimprovements and modifications can be practiced within the scope of theappended claims.

REFERENCES

The pertinent portions of the following references are incorporatedherein by reference.

-   1. Cyclized Cembranoids of Natural Occurrence Wahlberg, I.; Eklund,    A.-M. Prog. Chem. Org. Nat. Prod. 1992, 60, 1-141; Survey of    Oxygenated 2,11-Cyclized Cembranoids of Marine Origin Bernardelli,    P.; Paquette, L. A. Heterocycles 1998, 49, 531-556.-   2. Sclerophytins A and B. Isolation and Structures of Novel    Cytotoxic Diterpenes from the Marine Coral Sclerophytum capitalis    Sharma, P.; Alam, M. J. Chem. Soc. Perkin Trans 11988, 2537-2540.-   3. Structure revision: Revised Constitution of Sclerophytins A and B    Friedrich, D.; Doskotch, R. W.; Paquette, L. A. Org. Lett. 2000, 2,    1879-1882; Structural and stereochemical reassessment of    sclerophytin-type diterpenes Friedrich, D.; Paquette, L. A. J. Nat.    Prod. 2002, 65, 126-130.-   4. Total syntheses: Total Asymmetric Synthesis of the Putative    Structure of the Cytotoxic Diterpenoid (−)-Sclerophytin A and of the    Authentic Natural Sclerophytins A and B Bernardelli, P.; Moradei, O.    M.; Friedrich, D.; Yang, J.; Gallou, F.; Dyck, B. P.; Doskotch, R.    W.; Lange, T.; Paquette, L. A. J. Am. Chem. Soc. 2001, 123,    9021-9032; A General Strategy for the Synthesis of Cladiellin    Diterpenes Enantioselective Total Syntheses of    6-Acetoxycladiell-7(16), 1′-dien-3-ol (Deacetoxyalcyonin Acetate),    Cladiell-1′-ene-3,6,7-triol, Sclerophytin A, and the Initially    Purported Structure of Sclerophytin A MacMillan, D. W. C.;    Overman, L. E.; Pennington, L. D. Journal of the American Chemical    Society 2001, 123, 9033-9044. See also, A General Strategy for    Synthesis of Both (6Z)-and (6E)-Cladiellin Diterpenes: Total    Syntheses of (−)-Cladiella-6,11-dien-3-ol, (+)-Polyanthellin A,    (−)-Cladiell-11-ene-3,6,7-triol, and (−)Deacetoxyalcyonin Acetate.    Kim, H.; Lee, H.; Kim, J.; Kim, S.; Kim, D. J. Am. Chem. Soc. 2006,    128, 15851-15855.-   5. Eleutherobin, a New Cytotoxin that Mimics Paclitaxel (Taxol) by    Stabilizing Microtubules Lindel, T.; Jensen, P. R.; Fenical, W.;    Long, B. H.; Casazza, A. M.; Carboni, J.; Fairchild, C. R. J. Am.    Chem. Soc. 1997, 119, 8744-8745.-   6. Total syntheses: Total Synthesis of Eleutherobin and Eleuthosides    A and B Nicolaou, K. C.; Ohshima, T.; Hosokawa, S.; van Delft, F.    L.; Vourloumis, D.; Xu, J. Y.; Pfefferkorn, J.; Kim, S. J. Am. Chem.    Soc. 1998, 120, 8674-8680; The Total Synthesis of Eleutherobin Chen,    X.-T.; Bhattacharya, S. K.; Zhou, B.; Gutteridge, C. E.;    Pettus, T. R. R.; Danishefsky, S. J. J. Am. Chem. Soc. 1999, 121,    6563-6579.-   7. Sarcodictyin A and sarcodictyin B, novel diterpenoidic alcohols    esterified by (E)-N(1)-methylurocanic acid. Isolation from the    Mediterranean stolonifer Sarcodictyon roseumD'Ambrosio, M.;    Guerriero, A.; Pietra, F. Helv. Chim. Acta 1987, 70, 2019-2027;    Isolation from the Mediterranean stoloniferan coral Sarcodictyon    roseum of sarcodictyin C, D, E, and F, novel diterpenoidic alcohols    esterified by (E)-or(Z)-N(1)-methylurocanic acid. Failure of the    carbon-skeleton type as a classification criterion D'Ambrosio, M.;    Guerriero, A.; Pietra, F. Helv. Chim. Acta 1988, 71, 964-976.-   8. Total synthesis: Total Synthesis of Sarcodictyins A and B    Nicolaou, K. C.; Xu, J. Y.; Kim, S.; Pfefferkorn, J.; Ohshima, T.;    Vourloumis, D.; Hosokawa, S. J. Am. Chem. Soc. 1998, 120, 8661-8673;-   9. Cell-based screen for antimitotic agents and identification of    analogues of rhizoxin, eleutherobin, and paclitaxel in natural    extracts Roberge, M.; Cinel, B.; Anderson, H. J.; Lim, L.; Jiang,    X.; Xu, L.; Bigg, C. M.; Kelly, M. T.; Andersen, R. J. Cancer Res.    2000, 60, 5052-5058.-   10. Sarcodictyins: a New Class of Marine Derivatives with Mode of    Action Similar to TaxolCiomei, M.; Albanese, C.; Pastori, W.;    Grandi, M.; Pietra, F.; D'Ambrosio, M.; Guerriero, A.;    Battistini, C. Proc. Am. Assoc. Cancer Res. 1998, 38, 30; The    Coral-derived Natural Products Eleutherobin and Sarcodictyins A and    B: Effects on the Assembly of Purified Tubulin with and without    Microtubule-associated Proteins and Binding at the Polymer Taxoid    Site. Hamel, E.; Sackett, D. L.; Vourloumis, D.; Nicolaou, K. C.    Biochemistry 1999, 38, 5490-5498.-   11. Solid and Solution Phase Synthesis and Biological Evaluation of    Combinatorial Sarcodictyin Libraries Nicolaou, K. C.; Winssinger,    N.; Vourloumis, D.; Ohshima, T.; Kim, S.; Pfefferkorn, J.; Xu, J.    Y.; Li, T. J. Am. Chem. Soc. 1998, 120, 10814-10826.-   12. Synthesis of novel simplified sarcodictyin/eleutherobin analogs    with potent microtubule-stabilizing activity, using ring closing    metathesis as the key-step Beumer, R.; Bayon, P.; Bugada, P.; Ducki,    S.; Mongelli, N.; Sirtori, F. R.; Telser, J.; Gennari, C.    Tetrahedron 2003, 59, 8803-8820.-   13. Structure-activity Profiles of Eleutherobin Analogs and their    Cross-resistance in Taxol-resistant Cell Lines McDaid, H. M.;    Bhattacharya, S. K.; Chen, X. T.; He, L.; Shen, H. J.;    Gutteridge, C. E.; Horwitz, S. B.; Danishefsky, S. J. Cancer    Chemother. Pharmacol. 1999, 44, 131-137.-   14. Synthetic Transformations of Eleutherobin Reveal New Features of    Its Microtubule-Stabilizing Pharmacophore Britton, R.; Dilip de    Silva, E.; Bigg, C. M.; McHardy, L. M.; Roberge, M.;    Andersen, R. J. J. Am. Chem. Soc. 2001, 123, 8632-8633.-   15. Synthesis of a simplified analogue of eleutherobin via a Claisen    rearrangement and ring closing metathesis strategy Chiang, G. C. H.;    Bond, A. D.; Ayscough, A.; Pain, G.; Ducki, S.; Holmes, A. B. Chem.    Commun. 2005, 1860-1862.-   16. Design, synthesis and cytotoxic studies on the simplified oxy    analog of eleutherobinChandrasekhar, S.; Jagadeshwar, V.;    Narsihmulu, C.; Sarangapani, M.; Krishna, D. R.; Vidyasagar, J.;    Vijay, D.; Sastry, G. N. Bioorg. Med. Chem. Lett. 2004, 14,    3687-3689.-   17. See also, Synthetic Approach to Analogues of the Original    Structure of Sclerophytin A Jung, M. E.; Pontillo, J. J. Org. Chem.    2002, 67, 6848-6851.-   18. Davidson, J. E. P.; Gilmour, R.; Ducki, S.; Davies, J. E.;    Green, R.; Burton, J. W.; Holmes, A. B. Synlett 2004, 1434-1435; see    also, Chiang, G. C. H.; Bond, A. D.; Ayscough, A.; Pain, G.; Ducki,    S.; Holmes, A. B. Chem. Commun. 2005, 1860-1862.-   19. (a) A Novel Cycloaldol Approach to the Isobenzofuran Core of the    Eunicellin DiterpenesChai, Y.; Vicic, D. A.; McIntosh, M. C. Org.    Lett. 2003, 7, 1039-1042. (b) Studies directed toward the synthesis    of the massileunicellins Chai, Y.; McIntosh, M. C. Tetrahedron Lett.    2004, 45, 3269-3272. (c) Hutchison, J. M.; Lindsay, H. A.; Dormi, S.    S.; Jones, G. D.; Vicic, D. A.; McIntosh, M. C. Org. Lett. 2006, 8,    3663-3665.-   20. Direct Oxidation of Tertiary Allylic Alcohols. A Simple and    Effective Method for Alkylative Carbonyl Transposition Dauben, W.    G.; Michno, D. M. J. Org. Chem. 1977, 42, 682-685.-   21. Benzyl Enol Ethers via Decarboxylation of a-Benzyloxy-b-lactones    Derived from the Lithium a-Benzyloxy-a-lithioacetate Synthon Adam,    W.; Arias Encarnacion, L. A. Synthesis 1979, 388-390.-   22. Rapid colorimetric assay for cellular growth and survival:    application to proliferation and cytotoxicity assays. Mosmann T. J.    Immunol. Methods 1983, 65, 55-63.-   23. Modulation of mitogen-activated protein kinases and    phosphorylation of Bcl-2 by vinblastine represent persistent forms    of normal fluctuations at G2-M1. Fan, M.; Du, L.; Stone, A. A.;    Gilbert K. M.; Chambers, T. C. Cancer Res. 2000, 60, 6403-6407.-   24. Feasibility of drug screening with panels of human tumor cell    lines using a microculture tetrazolium assay. Alley, M. C.;    Scudiero, D. A.; Monks A.; Hursey, M. L.; Czerwinski, M. J.;    Fine, D. L.; Abbott, B. J.; Mayo, J. G.; Shoemaker, R. H.;    Boyd M. R. Cancer Res. 1988, 48, 589-601.

What is claimed is:
 1. A compound having a structural formula selectedfrom the following:

wherein R represents 14, substituted lower alkyl, or unsubstituted loweralkyl; Ar represents a substituted or unsubstituted aryl; andpharmaceutically acceptable esters and salts thereof.
 2. A compound ofclaim 1, wherein R=methyl, ethyl, cyclopropylmethyl orcyclopentylmethyl.
 3. A compound of claim 1, wherein Ar=2-Br-phenyl,3-Br-phenyl, 2,3-di-Cl-phenyl, 2,4-di-Cl-phenyl, 1-naphthyl, 2-pyridyl,2-furyl, or 2-fluorophenyl.
 4. A compound of claim 1 having structure10, wherein Ar=2-fluorophenyl.
 5. A method of converting (S)-(+)-carvoneto an aryl glycolate compound thereof, comprising: (i) reacting(S)-(+)-carvone with an arylaldehyde in an aldol condensation reactionto afford an aryl anti-alcohol; and (ii) etherifying the arylanti-alcohol to afford said aryl glycolate compound.
 6. The method ofclaim 5, further comprising cyclizing said aryl glycolate to afford anisobenzofuran having the structural formula

wherein R represents H, substituted lower alkyl, or unsubstituted loweralkyl; and Ar represents a substituted or unsubstituted aryl.
 7. Themethod of claim 6, further comprising converting by oxidativerearrangement said isobenzofuran to an enone having the structuralformula

wherein R represents H, substituted lower alkyl, or unsubstituted loweralkyl; and Ar represents a substituted or unsubstituted aryl.
 8. Themethod of claim 5, further comprising converting byβ-lactonization-decarboxylation said aryl glycolate to a diene havingthe structural formula

wherein Ar represents a substituted or unsubstituted aryl.
 9. A methodof inhibiting cell growth comprising contacting cancerous cells with agrowth inhibiting amount of a compound of claim 1, wherein the cancerouscells comprise leukemia, prostate, or non-small cell lung cancer cells.10. A method of inhibiting cell proliferation in a patient comprisingadministering to the patient a proliferation-inhibiting amount of acompound of claim 1, wherein the patient suffers from cancer comprisingleukemia, prostate, or non-small cell lung cancer.